Enzymimmunologischer Nachweis von ''pregnancy-associated glycoprotein'' (PAG) als Hilfsmittel zur Trächtigkeitsdiagnose beim Rind (English shop)

Morten Friedrich (Author)

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During pregnancy in cattle, various substances are synthesised in the placenta and released to the dam via complex mechanisms. These substances include a group of acidic glycoproteins, the “pregnancy-associated glycoproteins” (PAGs), which are produced in such large quantities that they are detectable in the blood and also in the milk of the dam and can be used as an indicator of the presence of pregnancy.

The aim of this study was to establish a quantitative enzyme immunoassay for bovine PAG-1 and to examine whether and from what point in time its detection can be regarded as a reliable sign of pregnancy. The enzyme immunological procedure was a competitive assay based on the double-antibody method (ELISA). The development of the detection method was divided into the following steps:

• Establishment of an ELISA for the determination of PAG in blood
• Adaptation of the ELISA for the detection of PAG in milk
• Compilation of PAG profiles in blood and milk of pregnant cows
• Testing of the PAG ELISA for its suitability for pregnancy diagnosis.

The establishment of a specific PAG ELISA was intended to create an alternative to the radioimmunological detection methods (RIA) used to date. The PAG antisera used in the ELISA, as well as the purified PAG required for the production of a biotin-labelled tracer, were identical to the substances used in the RIA and were provided to us by Prof. Beckers (ULG, Belgium). The correlation between ELISA and RIA was determined on the basis of 187 samples and was r=0.96 (p<0.01). Coefficients of variation (CV) and recovery rates (RR) of the ELISA were calculated for the measuring range from 0.4 to 12.5 ng/ml, according to which the ELISA is suitable for detecting minimal PAG concentrations in a milk or serum sample during pregnancy. Depending on the concentration, the intra-assay CV of the ELISA was 2–14% for blood serum and 10–12% for skimmed milk; the corresponding inter-assay CV was between 8–22% (serum) and 12–22% (milk). The lower detection limit of both assays was 0.4 ng/ml. To depict PAG profiles over the course of pregnancy, blood and milk samples were taken at regular intervals from 10 pregnant cows and their PAG content was determined by ELISA. The PAG profiles obtained showed the typical course described in the literature, with an initially stepwise increasing concentration during early pregnancy and a massive rise in the last days before parturition. The concentration in milk showed a significant correlation (r=0.81; p<0.01) with the values measured in blood, but overall ran at a considerably lower level, amounting to only between 3 and 8% of the serum concentration determined, according to which detection of pregnancy on the basis of a milk sample is not possible before day 150. Once development of the assay had been completed, it was to be investigated from what point in time and with what reliability the PAG ELISA can be used as a pregnancy test in cattle. For this purpose, a blood sample was taken from 400 cows between days 20 and 50 after insemination and analysed by ELISA. The pregnancy status of the animals was subsequently determined by rectal or ultrasonographic examination or by a further PAG sample. Daily and weekly mean values of the animals examined were calculated for the measured PAG concentrations. A significant difference (p<0.01; Dunnett’s t-test) in the PAG concentration of pregnant and non-pregnant animals was demonstrated from day 23, and a highly significant difference (p<0.001; Dunnett’s t-test) from day 28 after insemination. After day 30 following service, 94% of all animals, and from day 40 even 99%, were correctly identified as pregnant or non-pregnant by means of the PAG test. A possible explanation for incorrect diagnoses could be the occurrence of embryonic death between the day of the PAG analysis and the later pregnancy examination of the animals. In summary, it can be stated that the bPAG-1 ELISA represents a safe and reliable means of measuring the PAG concentration of a milk or serum sample and permits a pregnancy diagnosis in cows and heifers from day 30 after insemination on the basis of a serum sample.

ISBN-13 (Printausgabe)	3867270465
ISBN-13 (Hard Copy)	9783867270465
ISBN-13 (eBook)	9783736920460
Final Book Format	A5
Language	German
Page Number	158
Lamination of Cover	matt
Edition	1
Volume	0
Publication Place	Göttingen
Place of Dissertation	Göttingen
Publication Date	2006-11-06
General Categorization	Dissertation
Departments	Agricultural science